Kidney Week

Abstract: TH-PO0116

Proximity Labeling Proteomics Identifies Epigenetic and Chromatin Regulatory Complexes Associated with WT1

Session Information

• Integrative Genomics and Multi-Omics in Noncystic Genetic Kidney Disease
  October 22, 2026 | Location: Exhibit Hall A, Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Genetic Diseases of the Kidneys

• 1202 Genetic Diseases of the Kidneys: Non-Cystic (Complex and Non-Cystic Monogenic)

Authors

• Bilgin, Feride Melisa, Nationwide Children's Hospital Department of Nephrology, Columbus, Ohio, United States

Feride Melisa Bilgin, MS, MSc

• Albalawy, Wafaa N., Nationwide Children's Hospital Department of Nephrology, Columbus, Ohio, United States

Wafaa N. Albalawy, MS, PhD

• Wang, Xin, Nationwide Children's Hospital Department of Nephrology, Columbus, Ohio, United States

Xin Wang, PhD

• Bolsius, Leah, Boston Children's Hospital, Department of Nephrology, Boston, Massachusetts, United States

Leah Bolsius, MD

• Nelson, Becca, Boston Children's Hospital, Department of Nephrology, Boston, Massachusetts, United States

Becca Nelson

• Hiltebeitel, Lily, Boston Children's Hospital, Department of Nephrology, Boston, Massachusetts, United States

Lily Hiltebeitel

• Hildebrandt, Friedhelm, Boston Children's Hospital, Department of Nephrology, Boston, Massachusetts, United States

Friedhelm Hildebrandt, MD

• Mann, Nina, Nationwide Children's Hospital Department of Nephrology, Columbus, Ohio, United States

Nina Mann, MD

Background

The transcription factor Wilms' Tumor 1 (WT1) plays essential roles in nephrogenesis and the maintenance of mature podocytes. Pathogenic variants in WT1 cause a severe, progressive podocytopathy marked by heavy proteinuria and progression to kidney failure. Over 36 different WT1 isoforms exist, including WT1 (-)KTS and WT1 (+)KTS. Although WT1 regulates the expression of key podocyte proteins, the upstream mechanisms that govern WT1 function itself are poorly understood. Traditional methods for identifying protein-protein interactions, such as affinity purification approaches, are poorly suited for WT1, which is difficult to solubilize. Thus, we applied a proximity labeling approach to identify WT1-interacting proteins, aiming to better understand the factors that regulate WT1 function.

Methods

We generated stable hsPodocyte cell lines expressing wild type WT1 (-)KTS, mutant WT1 (-)KTS (p.Arg467Trp and p.Val167Asp), or wild type WT1 (+)KTS proteins conjugated to miniTurbo. Following biotin labeling, WT1-proximity proteins were enriched using streptavidin pulldown and identified by label-free mass spectrometry. Proximal proteins were analyzed using pathway enrichment analyses included GO and KEGG analyses. Select candidate interactions were validated by co-immunoprecipitation.

Results

We identified several chromatin-associated regulatory complexes in proximity to WT1, including components of the COMPASS complex, a histone H3K4 methyltransferase. Physical interaction between WT1 and the histone demethylase KDM6A was confirmed by co-immunoprecipitation. Pathway analyses revealed that proteins involved in double-strand DNA repair and stem cell maintenance were enriched in proximity to the wild-type WT1 (-)KTS protein, and that this was abrogated by the p.Arg467Trp mutation. Finally, despite their distinct functions, we found that the WT1 (-)KTS and WT1 (+)KTS isoforms exhibited significant overlap in their proximal proteomes.

Conclusion

By applying a proximity labeling approach, we identified epigenetic and chromatin regulatory complexes associated with WT1 in a human immortalized podocyte cell line and identified differential proximity proteomes between wild type and mutant WT1 proteins. Our results provide insight into the mechanisms by which WT1 transcriptional activity may be modulated.